Glia exchange molecules with axons in crayfish nervous tissue but the functional significance of such transfers has not been demonstrated. Defects of glia-axon exchanges in humans could contribute to the pathology of multiple sclerosis or convulsive disorders, where glial function is impaired. The long-term goal of this research is to determine whether glial cells contribute glutamic acid decarboxylase (GAD), or its product GABA, to axons in crayfish peripheral nerves. Following separation from their cell bodies, the motor axons in these nerves exhibit few ultrastructural or electrophysiological changes for very long times; also, neurotransmission and GAD levels are maintained. Published data suggests, indirectly, that GAD in these nerves is synthesized by glia surrounding inhibitory motor axons and that these glia might sustain inhibitory neurotransmission. This pilot project seeks to develop a technique for specific labeling of GAD in crayfish nerves. It is based upon a procedure which successfully labels GAD in mammalian brain. The rationale is that radioactive Gamma-acetylenic GABA inactivates GAD irreversibly in homogenates of nervous tissue after non-specific binding sites (e.g. GABA transaminase, GABA-T) have been titrated with gabaculine. The effects of these 2 drugs on GAD and GABA-T activities in homogenates of crayfish nerve tissues will be measured, using sensitive radiochemical assay procedures. (Alpha-H3)-acetylenic GABA will be synthesized. The specificity of binding of this radioactive ligand to GAD in the homogenates will be determined by correlating the amount of ligand bound with the degree of inactivation of GAD. The specificity of binding in the intact tissues then will be studied. First, the binding of the compound in intact tissue and an homogenate of an equivalent piece of tissue will be compared. Then changes in tissue GAD activity will be compared with changes in binding of the radioactive ligand in vivo. Finally, experiments will be performed to determine whether H3-acetylenic GABA penetrates neurons and glial cells in crayfish nerves. If the procedure permits specific labeling of GAD in axons and glia of crayfish peripheral nerves, then it will be employed subsequently to examine the cellular distribution of GAD